This work aims to identify the molecular mechanisms of cold detection, using the fruit fly Drosophila melanogaster as a model. Sensory systems play key roles in human health. Not only is human sensory system dysfunction associated with disease, the detection of temperature is important for the survival and host-seeking behavior of insect vectors of human disease. At present, our molecular understanding of temperature detection in insects is limited: insect cold receptors have not been reported. Our preliminary data suggest two Ionotropic Receptor family members, IR21a and IR25a, are involved in cold sensing. Ionotropic receptors (IRs) are a subfamily of iGluRs in invertebrates (Drosophila has 66 IRs) known to act not as mediators of glutamate- dependent synaptic transmission, but as chemoreceptors for acids and amines. We are proposing to test the hypothesis that specific isoforms of IR21a and IR25a drive cold avoidance by acting in the cold sensors of the arista and/or labellum. Using cell-specific rescue experiments, we propose to determine which IR21a and IR25a isoforms mediate cold avoidance, and to test whether expression of these IRs in cold sensors is necessary and/or required for cold avoidance. We will also test if IR21a and IR25a mediate cold sensing. Using a combination of calcium imaging and electrophysiology, we will examine the contribution of IR21a and IR25a to cold sensing by examining how the cold-responsiveness of each set of cold sensors is altered in mutants lacking these proteins. I will also test whether IR21a and IR25a act forming a cold receptor by ectopic expression of these proteins in the fly and by heterologous expression in frog oocytes and in cultured mammalian cells. These studies will contribute for the understanding of cold-reception and the identification of the receptors will be extremely significant to elucidate the molecular mechanisms involved.